Cloning and characterization of murine carnitine acetyltransferase: evidence for a requirement during cell cycle progression

Abstract

We have employed a newly developed differential screening technique (reverse strand priming) to identify murine carnitine acetyltransferase (CARAT) as a growth- and cell cycle-regulated gene in S3T3 mouse fibroblasts. Sequence analysis of the full-length cDNA clone and homology comparisons have revealed 87% homology to the human CARAT gene. On Northern blots we were able to measure a 2-3-fold induction 18 h after a mitogenic stimulus following serum deprivation as well as after release from a sodium butyrate block. The cell cycle induction pattern of the CARAT gene was analysed in mouse fibroblasts at different stages of the unperturbed cell cycle. Fractions obtained by elutriation of an exponentially growing culture showed a biphasic maximum of transcript abundance in the G1 and G2 phases of the cell cycle. CARAT expression was investigated in several organs of the adult mouse. Among those measured, CARAT expression was highest, relative to liver, in heart muscle (56-fold) and testis (21-fold). Using both conventional antisense oligodeoxynucleotides and novel single-stranded antisense phagemid DNA, we obtained evidence that the CARAT enzyme function is necessary for progression through G1 and into the S-phase of the cell cycle.